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作者 Stella S, Vitale SR, Martorana F, Massimino M, Pavone G, Lanzafame K, Bianca S, Barone C, Gorgone C, Fichera M, Manzella L
Stefania Stella, 1,2 Silvia Rita Vitale, 1,2 Federica Martorana, 1,2 Michele Massimino, 1,2 Giuliana Pavone, 3 Katia Lanzafame, 3 Sebastiano Bianca, 4 Chiara Barone, 5 Cristina Gorgone, 6 Marco Ficheraria, 1 Maperimental Clinic, 1 Maperimental Clinic, Maperimental Clinic University of Catania, Catania, 95123, Italy;2 Center for Experimental Oncology and Hematology, AOU Policlinico "G.Rodolico - San Marco", Catania , 95123, Italy; 3 Medical Oncology, AOU Policlinico "G. Rodolico - San Marco", Catania, 95123, Italy; 4 Medical Genetics, ARNAS Garibaldi, Catania, 95123, Italy; 5 Mankhwala Genetics, ASP, Syracuse, 96100, Italy; 6 Dipatimenti ya Sayansi ya Zamankhwala ndi Zamoyo, Yunivesite ya Catania, Zachibadwa Zamankhwala, Catania, Italy, 95123; 7Oasi Research Institute-IRCCS, Troina, 94018, Italy Kulankhulana: Stefania Stella, tel +39 095 378 1946, imelo [email protected]; [email protected] Cholinga: Kusintha kwa majeremusi mu BRCA1 ndi BRCA2 ndi kukhazikitsa khansa ya m'mawere (BC), ovary (OC) ndi zina zomwe zimagwirizanitsidwa ndi chiopsezo cha khansa cha moyo wonse. Kuyesa jini ya BRCA ndikofunikira kwambiri poyesa chiopsezo cha munthu aliyense, komanso kupeza njira zopewera matenda athanzi komanso njira zochiritsira odwala khansa. Kuchuluka kwa kusintha kwa BRCA1 ndi BRCA2 kumasiyana kwambiri m'madera osiyanasiyana, ndipo ngakhale kuti pali deta pa mitundu yosiyanasiyana ya matenda a BRCA m'mabanja a ku Sicily, maphunziro omwe amayang'ana makamaka anthu kum'mawa kwa Sicily akusowa. Cholinga cha kafukufuku wathu chinali kufufuza kuchuluka ndi kufalikira kwa kusintha kwa majeremusi a BRCA m'gulu la odwala a BC ochokera kum'mawa kwa Sicily ndikuwunika kuyanjana kwawo ndi makhalidwe enaake a BC pogwiritsa ntchito kutsatizana kwa m'badwo wotsatira. Kupezeka kwa kusintha komwe kumagwirizana ndi kuchuluka kwa chotupa ndi index ya kuchuluka.ZOTSATIRA: Ponseponse, odwala 35 (9%) anali ndi mtundu wa BRCA pathogenic, 17 (49%) mu BRCA1 ndi 18 (51%) mu BRCA2. Kusintha kwa BRCA1 kuli ponseponse mwa odwala a BC omwe ali ndi kachilombo katatu, pomwe kusintha kwa BRCA2 kuli ponseponse mwa odwala a BC omwe ali ndi kachilombo katatu. Poyerekeza ndi anthu omwe si odwala, anthu omwe ali ndi mitundu yosiyanasiyana ya BRCA1 anali ndi kalasi yapamwamba kwambiri ya chotupa komanso kuchuluka kwa khansa. Mapeto: Zomwe tapeza zikupereka chithunzithunzi cha momwe BRCA imasinthira m'thupi mwa odwala a BC ochokera kum'mawa kwa Sicily ndipo zimatsimikizira udindo wa kusanthula kwa NGS pozindikira odwala omwe ali ndi BC yobadwa nayo. Ponseponse, deta iyi ikugwirizana ndi umboni wakale womwe umathandizira kuyezetsa BRCA kuti apewe komanso kuchiza khansa moyenera mwa odwala omwe ali ndi kusintha kwa thupi.
Khansa ya m'mawere (BC) ndi khansa yofala kwambiri padziko lonse lapansi komanso khansa yoopsa kwambiri mwa akazi.1 Zizindikiro zamoyo zomwe zimatsimikizira kuti BC ikupita patsogolo komanso momwe imakhalira yaphunziridwa kwambiri ndipo zafotokozedwa pang'ono pakapita nthawi. Ndipotu, zizindikiro zingapo zotsatizana zimagwiritsidwa ntchito pogawa BC m'magulu osiyanasiyana a maselo. Ndi estrogen (ER) ndi/kapena progesterone receptor (PgR), human epidermal growth factor receptor 2 (HER2) amplification, proliferation index Ki-67 ndi tumor grade (G).2 Kuphatikiza kwa zinthuzi kunazindikira magulu otsatirawa a BC: 1) Luminal tumors, zomwe zikuwonetsa ER ndi/kapena PgR expression, zinali 75% ya BCs. Zotupa izi zinagawidwanso kukhala Luminal A, pomwe Ki-67 inali pansi pa 20% ndi HER2 negative, ndi Luminal B, pomwe Ki-67 inali yofanana kapena yoposa 20% komanso pamaso pa HER2 amplification, mosasamala kanthu za proliferation index; 2) Zotupa za HER2+ zomwe zili ndi ER ndi PgR negative koma zikuwonetsa HER2 amplification. Gululi limawerengera 10% ya zotupa zonse za m'mawere; 3) Khansa ya m'mawere yopanda katatu (TNBC), yomwe sikuwonetsa ER ndi PgR expression ndi HER2 amplification, imawerengera pafupifupi 15% ya khansa ya m'mawere.2-4
Pakati pa mitundu iyi ya BC, kuchuluka kwa chotupa ndi kuchuluka kwa chotupa zimayimira zizindikiro zoyambira zomwe zimagwirizana mwachindunji komanso paokha ndi kuopsa kwa chotupa komanso kupezeka kwa matendawa.5,6
Kuwonjezera pa zinthu zamoyo zomwe zatchulidwa pamwambapa, udindo wa kusintha kwa majini komwe kumatsogolera ku kukula kwa BC wakhala wofunika kwambiri m'zaka zingapo zapitazi.7 Pafupifupi 1 mwa 10 zotupa za m'mawere zimabadwa chifukwa cha kusintha kwa majeremusi m'majini enaake.8 Kafukufuku awiri akuluakulu okhudza matenda opatsirana omwe amakhudza akazi oposa 180,000 posachedwapa adapeza gulu la majini asanu ndi atatu (monga ATM, BARD1, BRCA1, BRCA2, CHK2, PALB2, RAD51C, ndi RAD51D) omwe makamaka amachititsa BC yobadwa nayo. Pakati pa majini awa, BRCA1 ndi BRCA2 (yomwe tsopano ikutchedwa BRCA1/2) yawonetsa mgwirizano wamphamvu kwambiri ndi kukula kwa zotupa za m'mawere.9-12 Ndipotu, kusintha kwa majeremusi a BRCA1/2 kumawonjezera kwambiri chiopsezo cha moyo wonse cha BC komanso khansa zina, kuphatikizapo mazira, prostate, pancreatic, colorectal, ndi melanoma. Kuyambira zaka 13 mpaka 80, kuchuluka kwa BC ndi 72% mwa akazi omwe ali ndi BRCA1 pathogenic variant (PV) ndi 69% mwa akazi omwe ali ndi BRCA2 PV.14
Chochititsa chidwi n'chakuti, buku laposachedwapa likusonyeza kuti chiopsezo cha BC chimadalira mtundu wa PV. Ndipotu, poyerekeza ndi mitundu yotupa ya tizilombo toyambitsa matenda, mitundu yolakwika yowonekera bwino, makamaka mu jini la BRCA1, imagwirizanitsidwa ndi chiopsezo chocheperako cha BC, makamaka mwa akazi okalamba.15
Kupezeka kwa BRCA1 kapena BRCA2 PV kunagwirizanitsidwa ndi zinthu zosiyanasiyana zamoyo ndi zachipatala.16,17 Ma BC ogwirizana ndi BRCA1 nthawi zambiri amakhala oopsa kwambiri, osasiyana bwino, komanso amakula kwambiri. Zotupa izi nthawi zambiri zimakhala ndi zotsatirapo zitatu ndipo zimakhala ndi unyamata woyambirira. Zotupa zomwe zimapezeka mwa odwala omwe asintha BRCA2 nthawi zambiri zimakhala ndi magiredi apakati mpaka osiyana bwino komanso zizindikiro zosinthika zofalikira. Zotupa izi zimapezeka kwambiri mu lumen B ndipo nthawi zambiri zimapezeka mwa okalamba.16-18 Chodziwika bwino ndichakuti, kusintha kwa BRCA1 ndi BRCA2 kumawonjezera chidwi ndi mankhwala enaake, kuphatikiza mchere wa platinamu ndi mankhwala enaake monga poly(ADP-ribose) polymerase inhibitors (PARPi).19,20
M'zaka zingapo zapitazi, kukhazikitsidwa kwa njira yotsatizana ya m'badwo wotsatira (NGS) m'machitidwe azachipatala kwathandiza odwala ambiri a BC kuti ayesedwe mozama za matenda omwe ali ndi khansa, kuphatikizapo BRCA1/2.21 Nthawi yomweyo, matanthauzidwe ozikidwa pa mfundo zenizeni zokhudzana ndi mbiri ya banja, kuchuluka kwa anthu, ndi makhalidwe a kliniki kuti adziwe bwino anthu oyenerera kuyesedwa kwa BRCA1/2.22,23 Pachifukwa ichi, umboni ukuwonjezeka pakuwunika kwa BRCA1/2 m'magulu enaake, zomwe zikuwonetsa kusiyana m'madera osiyanasiyana.24–27 Ngakhale pali malipoti okhudza gulu la BC kumadzulo kwa Sicily, pali deta yochepa pakuwunika kwa BRCA1/2 m'magulu a kum'mawa kwa Sicily.28,29
Apa tikufotokoza zotsatira za kuyezetsa kwa germline BRCA1/2 mwa odwala a BC ochokera kum'mawa kwa Sicily, zomwe zikugwirizana kwambiri ndi kupezeka kwa kusintha kwa BRCA1 kapena BRCA2 ndi zizindikiro zazikulu za matenda a zotupa izi.
Kafukufuku wobwerezabwereza unachitika ku "Center for Experimental Oncology and Hematology" ku Policlinico Hospital.Rodolico - San Marco ku Catania. Kuyambira Januwale 2017 mpaka Marichi 2021, odwala 455 omwe ali ndi khansa ya m'mawere ndi yam'mimba, melanoma, pancreatic kapena prostate adatumizidwa ku labotale yathu yowunikira mamolekyu kuti akayesedwe majini a BRCA/2. Kafukufukuyu adachitika motsatira Chikalata cha Helsinki, ndipo onse omwe adachita nawo adapereka chilolezo cholembedwa asanafufuze mamolekyu.
Makhalidwe a histological ndi achilengedwe (ER, PgR, HER2 status, Ki-67, ndi giredi) ya BC adayesedwa pa biopsy yayikulu kapena zitsanzo za opaleshoni, poganizira zigawo za chotupa choopsa zokha. Kutengera ndi makhalidwe awa, ma BC adagawidwa motere: luminal A (ER+ ndi/kapena PgR+, HER2-, Ki-67<20%), luminal B (ER+ ndi/kapena PgR+, HER2-, Ki-67≥20%), luminal B-HER2+ (ER ndi/kapena PgR+, HER2+), HER2+ (ER ndi PgR-, HER2+) kapena triple negative (ER ndi PgR-, HER2-).
Asanayese momwe BRCA1 ndi BRCA2 zilili, gulu la akatswiri osiyanasiyana kuphatikizapo katswiri wa khansa, katswiri wa majini, ndi katswiri wa zamaganizo adachita upangiri wa majini a chotupa kwa wodwala aliyense kuti adziwe kupezeka kwa BRCA1 ndi/kapena BRCA1. kapena anthu omwe ali ndi chiopsezo chachikulu cha PV mu jini la BRCA2. Kusankha wodwala kunachitika motsatira malangizo a Italian Society of Medical Oncology (AIOM) ndi malangizo am'deralo a Sicilian.30,31 Izi zikuphatikizapo: (i) mbiri ya banja ya mitundu yodziwika bwino ya matenda m'majini othawirako (monga BRCA1, BRCA2, TP53, PTEN); (ii) amuna omwe ali ndi BC; (iii) omwe ali ndi BC ndi OC; (iv) akazi omwe ali ndi BC <36 zaka, TNBC <60 zaka, kapena awiriawiri BC <50 zaka; (v) mbiri yachipatala ya BC <50 zaka ndi wachibale mmodzi wosachepera digiri yoyamba: (a) BC <50 zaka; (b) OC yosakhala ya mucinous komanso yopanda malire a msinkhu uliwonse; (c) awiriawiri BC; (d) amuna BC; (e) khansa ya kapamba; (f) khansa ya prostate; (vi) mbiri ya munthu wa BC wazaka zopitilira 50 ndi mbiri ya banja la BC, OC, kapena khansa ya kapamba kwa achibale omwe ali achibale oyamba kwa wina ndi mnzake (kuphatikiza achibale omwe ali achibale nawo oyamba); (vii) Mbiri ya munthu wa OC ndi wachibale mmodzi wosachepera woyamba: (a) BC pansi pa zaka 50; (b) NOC; (c) BC yapawiri; (d) BC yamwamuna; (vii) mkazi wokhala ndi OC yapamwamba kwambiri.
Chitsanzo cha magazi cha 20 mL chochokera kwa wodwala aliyense chinatengedwa ndi kutengedwa m'machubu a EDTA (BD Biosciences). DNA ya genomic inapezeka kuchokera ku zitsanzo za magazi athunthu a 0.7 mL pogwiritsa ntchito QIAsymphony DSP DNA Midi kit Isolation Kit (QIAGEN, Hilden, Italy) motsatira malangizo a wopanga ndipo inadutsa mu Qubit® 3.0 Fluorometer (Thermo Fisher Scientific, Waltham, MA, USA). Kukonzekera kwa cholinga ndi laibulale kumachitika ndi Oncomine™ BRCA Research Assay Chef, yokonzeka kuyikidwa mu Ion AmpliSeq™ Chef Reagents DL8 Kit kuti ikonzedwe yokha malinga ndi malangizo a wopanga. Chidachi chili ndi ma pool awiri a multiplex PCR primer omwe angagwiritsidwe ntchito kuphunzira majini onse a BRCA1 (NM_007300.3) ndi BRCA2 (NM_000059.3). Mwachidule, 15 µL ya DNA iliyonse yochepetsedwa (10 ng) idawonjezedwa pama plates okhala ndi barcode kuti ikonzedwe laibulale ndipo ma reagents onse ndi zinthu zina zogwiritsidwa ntchito zidayikidwa pa chida cha Ion Chef™. Kukonzekera kwa laibulale yokha ndi kusonkhanitsa zitsanzo za laibulale yokhala ndi barcode zidachitidwa pa chida cha Ion Chef™. Chiwerengero cha malaibulale okonzedwa kenako chidayesedwa ndi Qubit® 3.0 Fluorometer (Thermo Fisher Scientific, Waltham, MA, USA) malinga ndi malangizo a wopanga. Pomaliza, malaibulale amaphatikizidwa mu ma ratios ofanana m'machubu a zitsanzo za laibulale ya Ion Chef™. (machubu okhala ndi barcode) ndikuyikidwa pa chida cha Ion Chef™. Kutsata kunachitika pogwiritsa ntchito chida cha Ion Torrent S5 (Thermo Fisher Scientific) (Thermo Fisher Scientific) pogwiritsa ntchito Ion 510 Chip (Thermo Fisher Scientific). Kusanthula deta kunachitika ndi Amplicon Suite (SmartSeq srl) ndi Ion Reporter Software.
Mitundu yonse ya mayina inatsatira malangizo omwe alipo a Human Genome Variation Consortium, omwe akupezeka pa intaneti (HGVS, http://www.hgvs.org/mutnomen). Kufunika kwachipatala kwa mitundu ya BRCA1/2 kunatanthauzidwa pogwiritsa ntchito gulu la International Consortium ENIGMA (Evidence-Based Network for Interpreting Germline Mutant Alleles, https://enigmaconsortium.org/) ndi kufunsa ma database osiyanasiyana monga ARUP, BRCAEXCHANGE, ClinVar, IARC_LOVD, ndi UMD. Gululi limaphatikizapo magulu asanu osiyana a zoopsa: benign (gulu I), benign (gulu II), variant of uncertain significant (VUS, gulu III), benign pathogenic (gulu IV), ndi pathogenic (gulu V).VarSome adafufuzanso momwe masinthidwe amakhudzira kapangidwe ka mapuloteni ndi ntchito yake, chida chophunzitsira chomwe chili ndi mwayi wopeza ma database 30.32
Kuti tipereke kufunika kwa VUS iliyonse, njira zotsatirazi zowerengera mapuloteni zidagwiritsidwa ntchito: MUTATION TASTER, 33 PROVEAN-SIFT (http://provean.jcvi.org/index.php), POLYPHEN-2 (http:// /genetics.bwh.harvard.edu/pph2/) ndi Align-GVGD (http://agvgd.hci.utah.edu/agvgd_input.php). Zosintha zomwe zimagawidwa ngati kalasi 1 ndi 2 zimaonedwa ngati zamtundu wakuthengo.
Kusanthula kwa Sanger kunatsimikizira kupezeka kwa mtundu uliwonse wa matenda. Mwachidule, ma primer awiri apadera adapangidwa pa mtundu uliwonse wopezeka pogwiritsa ntchito ma gene reference sequences a BRCA1 ndi BRCA2 (NG_005905.2, NM_007294.3 ndi NG_012772.3, NM_000059.3, motsatana). Chifukwa chake, PCR yolunjika idachitidwa kutsatiridwa ndi Sanger sequencing.
Odwala omwe adapezeka kuti alibe jini la BRCA1/2 adayesedwa pogwiritsa ntchito multiplex ligation-dependent probe amplification (MLPA) motsatira malangizo a wopanga kuti awone ngati pali ma genomic rearrangements akuluakulu (LGR). Mwachidule, zitsanzo za DNA zimachotsedwa ndipo ma probe okwana 60 a BRCA1 ndi BRCA2 amagwiritsidwa ntchito, iliyonse ikupeza DNA sequence yeniyeni pafupifupi ma nucleotide 60 kutalika. Zinthu zokulitsa probe, zomwe zimakhala ndi ma PCR amplicons apadera, kenako zidawunikidwa ndi capillary electrophoresis ndi pulogalamu ya Cofalyser.Net mogwirizana ndi matebulo oyenera a Cofalyser (www.mrcholland.com).
Zosintha zina zosankhidwa za clinicopathological (histological grade ndi Ki-67% proliferation index) zinagwirizanitsidwa ndi kukhalapo kwa BRCA1/2 PV, zomwe zinawerengedwa pogwiritsa ntchito pulogalamu ya Prism v. 8.4 pogwiritsa ntchito mayeso enieni a Fisher poganiza kuti p-value <0.05 ndi yofunika kwambiri.
Pakati pa Januwale 2017 ndi Marichi 2021, odwala 455 adayesedwa ngati ali ndi ma germline BRCA1/2 mutations. Kuyesa kusintha kwa maselo kunachitika ku Policlinico Hospital's Center for Experimental Oncology and Hematology. Malinga ndi malangizo a Sicilian (http://www.gurs.regione.sicilia.it/Indicep1.htm, N. 02-Venerdì 10 Gennaio 2020), Rodolico wa ku Catania – San Marco” onse, odwala 389 anali ndi khansa ya m'mawere, khansa ya m'chiberekero 37, khansa ya kapamba 16, khansa ya prostate 8 ndi khansa ya melanoma 5. Kugawidwa kwa odwala malinga ndi mtundu wa khansa ndi zotsatira za kusanthula kwawonetsedwa mu Chithunzi 1.
Chithunzi 1 chikuwonetsa tchati cha momwe kafukufukuyu akuyendera chomwe chikuwonetsa mwachidule za kafukufukuyu. Odwala omwe ali ndi zotupa za m'mawere, melanoma, pancreatic, prostate, kapena ovarian adayesedwa kusintha kwa majini a BRCA1 ndi BRCA2.
Mafupikitsidwe: Ma PV, mtundu wa matenda; VUS, mtundu wosatsimikizika; WT, mtundu wachilengedwe wa BRCA1/2.
Tinasankha bwino maphunziro athu pa magulu a khansa ya m'mawere. Odwalawo anali ndi zaka zapakati pa 49 (zaka 23-89) ndipo ambiri anali akazi (n=376, kapena 97%).
Mwa anthuwa, 64 (17%) anali ndi kusintha kwa BRCA1/2 ndipo onse anali akazi. Makumi atatu ndi asanu (9%) anali ndi PV ndipo 29 (7.5%) anali ndi VUS. Khumi ndi asanu ndi awiri (48.6%) mwa mitundu 35 ya matenda inachitika mu BRCA1 ndi 18 (51.4%) mu BRCA2, pomwe 5 VUS idachitika mu BRCA1 (17.2%) ndi 24 (82.8%) mu BRCA2 (Zithunzi 1 ndi 2). LGR sinalipo mu kusanthula kwa MLPA.
Chithunzi 2. Kusanthula kwa kusintha kwa BRCA1 ndi BRCA2 mwa odwala khansa ya m'mawere 389. (A) Kufalikira kwa mitundu yosiyanasiyana ya matenda (PV) (ofiira), mitundu yosatsimikizika (VUS) (lalanje), ndi WT (buluu) mwa odwala khansa ya m'mawere 389; (B) odwala khansa ya m'mawere 389. Makumi atatu ndi asanu (9%) anali ndi mitundu yosiyanasiyana ya matenda a BRCA1/2 (PVs). Pakati pawo, 17 (48.6%) anali onyamula BRCA1 PV (ofiira wakuda) ndipo 18 (51.4%) anali onyamula BRCA2 (ofiira wopepuka); (C) 29 (7.5%) mwa anthu 389 omwe anali ndi VUS, majini 5 (17.2%) a BRCA1 (lalanje wakuda) ndi majini 24 (82.8%) a BRCA2 (lalanje wopepuka).
Mafupikitsidwe: Ma PV, mtundu wa matenda; VUS, mtundu wosatsimikizika; WT, mtundu wachilengedwe wa BRCA1/2.
Kenako tinafufuza kufalikira kwa ma subtypes a mamolekyu a BC mwa odwala omwe ali ndi BRCA1/2 PV. Kugawidwako kunaphatikizapo 2 (5.7%) luminal A, 15 (42.9%) luminal B, 3 (8.6%) luminal B-HER2+, 2 (5.7%) HER2+ ndi 13 (37.1%) odwala TNBC. Pakati pa odwala omwe ali ndi BRCA1, 5 (29.4%) anali ndi luminal B BC, 2 (11.8%) anali ndi matenda a HER2+, ndipo 10 (58.8%) anali ndi TNBC. Zotupa zopanda kusintha kwa BRCA1 zinali luminal A kapena luminal B-HER2+ (Chithunzi 3). Mu gulu laling'ono la BRCA2, zotupa 10 (55.6%) zinali luminal B, 3 (16.7%) zinali luminal B-HER2+, 3 (16.7%) TNBC ndi 2 (11.1%) zinali luminal A (Chithunzi 3). Ayi Zotupa za HER2+ zinalipo m'gululi. Chifukwa chake, kusintha kwa BRCA1 kumachitika kwambiri mwa odwala a TNBC, pomwe kusintha kwa BRCA2 kumachitika kwambiri mwa anthu omwe ali ndi lumen B.
Chithunzi 3 Kufalikira kwa mitundu yosiyanasiyana ya khansa ya m'mawere mwa odwala omwe ali ndi mitundu yosiyanasiyana ya BRCA1 ndi BRCA2. Ma histogram omwe akuwonetsa kufalikira kwa ma PV a BRCA1- (ofiira akuda) ndi BRCA2- (ofiira opepuka) pakati pa mitundu yosiyanasiyana ya odwala khansa ya m'mawere. Manambala omwe alembedwa mkati mwa bokosi lililonse akuyimira kuchuluka kwa odwala omwe ali ndi BRCA1 ndi BRCA2 PV pa mtundu uliwonse wa khansa ya m'mawere.
Mafupikitsidwe: Ma PV, mtundu wa matenda; HER2+, cholandirira kukula kwa munthu cha epidermal growth factor 2 chabwino; TNBC, khansa ya m'mawere ya katatu.
Pambuyo pake, tinayesa mtundu ndi malo a majini a BRCA1 ndi BRCA2 PVs. Mu BRCA1 PV, tinaona mitundu 7 ya nucleotide imodzi (SNVs), kuchotsedwa 6, kubwerezabwereza 3 ndi kuikidwa kamodzi. Kusintha kumodzi kokha (c.5522delG) kukuyimira kupeza kwatsopano. BRCA1 PV yodziwika kwambiri yomwe idapezeka mwa anthu onse awiri inali c.5035_5039delCTAAT. Kusinthaku kumaphatikizapo kuchotsedwa kwa ma nucleotide asanu (CTAAT) mu BRCA1 exon 15, zomwe zidapangitsa kuti amino acid leucine ilowe m'malo ndi tyrosine pa codon 1679, ndipo chifukwa cha kusintha kwa frameshift ndi codon yomwe idanenedweratu ipangitse kuti mapuloteni asamayende bwino. Kusintha kwina konse kumapezeka mu nkhani imodzi yokha. Chodziwika bwino n'chakuti, imodzi mwa ma PV omwe adanenedwa inali m'dera la mgwirizano wa malo a splice (c.4357+1G>T) (Table 1).
Ponena za BRCA2 PV, tawona kuchotsedwa 6, ma SNV 6 ndi kubwerezabwereza 2. Palibe kusintha komwe kwapezeka komwe ndi kwatsopano. Masinthidwe atatu adabweranso mwa anthu athu, c.428dup ndi c.8487+1G>A omwe adawonedwa mwa anthu atatu, kutsatiridwa ndi c.5851_5854delAGTT yomwe idapezeka m'magawo awiri. Kusintha kwa c.428dup kumaphatikizapo kubwereza kwa C mu exon 5 ya BRCA2, yomwe idanenedweratu kuti ipanga puloteni yodulidwa, yosagwira ntchito. Kusintha kwa c.8487+1G>A kumachitika m'chigawo cha intronic cha BRCA2 intron 19 (± 1,2) ndipo kumakhudza kutsatana kwa mgwirizano wa splicing, zomwe zimapangitsa kuti splicing isinthe zomwe zimapangitsa kuti puloteni ikhale yosazolowereka kapena yosakhalapo. Mtundu wa c.5851_5854delAGTT umachitika chifukwa cha kuchotsedwa kwa 4-nucleotide kuchokera ku malo a nucleotide 5851 mpaka 5854 mu exon 10 yolemba Jini la BRCA2 ndipo limabweretsa kusintha kwa frame ndi codon yoyimitsa yomwe yanenedweratu (p.S1951WfsTer). Chodziwika bwino, monga momwe zanenedwera kale, kusintha konse kwa c.631G>A ndi c.7008-2A>T kunapezeka mwa wodwala yemweyo.34 Kusintha koyamba kumaphatikizapo kusintha kwa adenosine (A) mu BRCA2 exon 7 ndi guanine (G) yokhala ndi nucleotide zomwe zimapangitsa kusintha kwa valine kukhala isoleucine pa codon 211, isoleucine Amino acid ndi amino acid yokhala ndi makhalidwe ofanana kwambiri. Kusinthaku kumakhudza kugawanika kwa mRNA kwabwinobwino. Mtundu wachiwiri uli m'dera la intronic ndipo umabweretsa kusintha kwa A kawiri kukhala thymine (T) isanafike exon 13 ya jini yomwe imalemba BRCA2. Kusintha kwa c.7008-2A>T kungapangitse zolemba zambiri zautali wosiyana. Kuphatikiza apo, m'gulu la BRCA2 PVs, zosintha 4 mwa 18 (22.2%) zinali intronic.
Kenako tinajambula masinthidwe olakwika a BRCA1/2 m'magawo ogwira ntchito komanso m'magawo omangira mapuloteni (Chithunzi 4). Mu jini ya BRCA1, 50% ya ma PV anali m'dera la khansa ya m'mawere (BCCR), pomwe 22% ya masinthidwewo anali m'dera la khansa ya ovarian (OCCR) (Chithunzi 4A). Mu BRCA2 PV, 35.7% ya mitundu yosiyanasiyana inali m'dera la BCCR ndipo 42.8% ya masinthidwewo anali mu OCCR (Chithunzi 4B). Kenako, tinayesa malo a PV mkati mwa madera a mapuloteni a BRCA1 ndi BRCA2. Pa puloteni ya BRCA1, tinapeza ma PV atatu m'malo ozungulira ndi ozungulira, ndi masinthidwe awiri m'malo a BRCT (Chithunzi 4A). Pa puloteni ya BRCA2, ma PV anayi adajambulidwa ku malo obwerezabwereza a BRC, pomwe kusintha kwa intronic ndi 3 kwa exonic kudapezeka m'malo a oligo/oligosaccharide-binding (OB) ndi tower (T) (Chithunzi 4B).
Chithunzi 4 Chiwonetsero cha ma protein a BRCA1 ndi BRCA2 ndi malo a mitundu yosiyanasiyana ya matenda. Chithunzichi chikuwonetsa kufalikira kwa mitundu yosiyanasiyana ya matenda a BRCA1 (A) ndi BRCA2 (B) mwa odwala khansa ya m'mawere. Kusintha kwa ma genetiki kumawonetsedwa mu buluu, pomwe mitundu yosiyanasiyana ya intronic imawonetsedwa mu lalanje. Kutalika kwa bar kumayimira chiwerengero cha milandu. Ma protein a BRCA1 ndi BRCA2 ndi madera awo ogwira ntchito anenedwa. (A) Puloteni ya BRCA1 ili ndi dera lozungulira (RING) ndi dongosolo la malo a nyukiliya (NLS), dera lozungulira, dera la SQ/TQ (SCD), ndi dera la C-terminal la BRCA1 (BRCT). (B) Puloteni ya BRCA2 ili ndi ma BRC repeats asanu ndi atatu, dera lolumikiza DNA lokhala ndi dera lozungulira (Helical), ma fold atatu a oligonucleotide/oligosaccharide-binding (OB), dera la nsanja (T), ndi An NLS kumbali ya C. Madera otchedwa Breast Cancer Cluster Region (BCCR) ndi Ovarian Cancer Cluster Region (OCCR) akuwonetsedwa pa pansi.* Zimayimira kusintha kwa majini komwe kumazindikira ma codon oima.
Kenako tinafufuza zizindikiro za BC clinicopathological zomwe zingagwirizane ndi kupezeka kwa BRCA1/2 PV. Zolemba zonse zachipatala zinalipo kwa odwala 181 omwe alibe BRCA1/2 (osanyamula) ndi onse omwe ali ndi matendawa (n = 35). Panali mgwirizano pakati pa kuchuluka kwa chotupa ndi giredi.
Tinawerengera kufalikira kwa Ki-67 kutengera apakati pa gulu lathu (25%, range <10-90%). Anthu omwe ali ndi Ki-67 <25% adatanthauzidwa kuti ndi "otsika Ki-67″, pomwe anthu omwe ali ndi mtengo wa ≥ 25% amaonedwa kuti ndi "okwera Ki-67″. Kusiyana kwakukulu kwa Ki-67 (p<0.01) kunapezeka pakati pa osakhala onyamula ndi onyamula BRCA1 PV (Chithunzi 5A).
Chithunzi 5 Kugwirizana kwa Ki-67 ndi kugawa kwa giredi mwa amayi omwe ali ndi BRCA1 ndi BRCA2 PVs ndi omwe alibe.(A) Chithunzi cha bokosi chomwe chikuwonetsa miyeso yapakati ya Ki-67 mwa odwala 181 omwe si onyamula BC poyerekeza ndi odwala BRCA1 (18) kapena BRCA2 (17) a PV. Miyeso ya P pansi pa 0.5 idawonedwa kuti ndi yofunika kwambiri pa ziwerengero.(B) Histogram yoyimira kugawa kwa odwala khansa ya BC m'magulu a giredi ya histological (G2 ndi G3) malinga ndi momwe BRCA1 ndi BRCA2 zilili (omwe ali ndi WT, onyamula BRCA1 ndi BRCA2 PVs).
Momwemonso, tinayang'ana ngati kuchuluka kwa chotupa kukugwirizana ndi kukhalapo kwa BRCA1/2 PV. Popeza G1 BC sinalipo mwa anthu athu, tinagawa odwala m'magulu awiri (G2 kapena G3). Mogwirizana ndi zotsatira za Ki-67, kusanthulaku kunavumbula mgwirizano wofunikira pakati pa kuchuluka kwa chotupa ndi kusintha kwa BRCA1, ndi kuchuluka kwakukulu kwa zotupa za G3 mwa omwe amanyamula BRCA1 poyerekeza ndi omwe sanyamula (p<0.005) (Chithunzi 5B).
Kupita patsogolo kwa ukadaulo wowerengera DNA kwathandiza kupita patsogolo kwambiri pa mayeso a majini a BRCA1/2, zomwe zimakhudza kwambiri odwala omwe ali ndi mbiri ya khansa m'banja lawo. Mpaka pano, mitundu pafupifupi 20,000 ya BRCA1/2 yapezeka ndikugawidwa m'magulu malinga ndi American Society of Medical Genetics 35 ndi dongosolo la ENIGMA.35,36 Ndizodziwika bwino kuti mawonekedwe a BRCA1/2 amasinthasintha kwambiri m'madera osiyanasiyana.37 Mu Italy, kuchuluka kwa ma PV a BRCA1/2 kunali kuyambira 8% mpaka 37%, kusonyeza kusiyana kwakukulu kwamkati mwa dziko.38,39 Ndi anthu pafupifupi 5 miliyoni, Sicily ndi dera lachisanu lalikulu kwambiri ku Italy pankhani ya chiwerengero cha anthu okhalamo. Ngakhale kuti pali deta yokhudza kufalikira kwa BRCA1/2 kumadzulo kwa Sicily, palibe umboni wokwanira kum'mawa kwa chilumbachi.
Kafukufuku wathu ndi umodzi mwa malipoti oyamba okhudza kuchuluka kwa BRCA1/2 PV mwa odwala a BC kum'mawa kwa Sicily.28 Tinayang'ana kwambiri kusanthula kwathu pa BC, chifukwa uwu ndiye matenda ofala kwambiri m'gulu lathu.
Poyesa odwala 389 BC, 9% anali ndi ma PV a BRCA1/2, omwe anagawidwa mofanana pakati pa BRCA1 ndi BRCA2. Zotsatirazi zikugwirizana ndi zomwe zidanenedwa kale mwa anthu aku Italy.28 Chochititsa chidwi n'chakuti, 3% (13/389) a gulu lathu anali amuna. Chiŵerengerochi ndi chachikulu kuposa momwe timayembekezera khansa ya m'mawere ya amuna (1% ya ma BC onse), 40 zomwe zikusonyeza kusankha kwathu kwa anthu kutengera chiopsezo cha kusintha kwa BRCA1/2. Komabe, palibe aliyense wa amunawa amene adapanga BRCA1/2 PV, kotero anali ofuna kufufuzidwa kwina kwa mamolekyu kuti athetse kupezeka kwa kusintha kosazolowereka monga PALB2, RAD51C ndi D, pakati pa ena. Zosintha zosatsimikizika zinapezeka mwa 7% ya anthu omwe BRCA2 VUS inali yoonekeratu. Ngakhale zotsatirazi zikugwirizana ndi umboni womwe ulipo kale.28,41,42
Pamene tidasanthula kufalikira kwa ma subtypes a mamolekyu a BC mwa akazi osinthika a BRCA1/2, tidatsimikiza mgwirizano wodziwika pakati pa TNBC ndi BRCA1 PV (58.8%) ndi pakati pa luminal B BC ndi BRCA2 PV (55.6%).16,43 Zotupa za luminal A ndi HER2+ mu BRCA1 ndi BRCA2 PV zonyamula zikugwirizana ndi zomwe zilipo kale.16,43
Kenako timayang'ana kwambiri mtundu ndi malo a BRCA1/2 PV. Mu gulu lathu, BRCA1 PV yodziwika kwambiri inali c.5035_5039delCTAAT. Ngakhale Incorvaia et al. sanafotokoze za mtundu uwu m'gulu lawo la ku Sicilian, olemba ena anena kuti ndi mtundu wa BRCA1 PV.34 Ma PV angapo a BRCA1 adapezeka m'gulu lathu - mwachitsanzo c.181T>G, c.514del, c.3253dupA ndi c.5266dupC - zomwe zawonedwa ku Sicily.28 Mwa izi, masinthidwe awiri oyambitsa BRCA1 (c.181T>G ndi c.5266dupC) amapezeka kwambiri mwa Ayuda a ku Ashkenazi aku Eastern ndi Central Europe (Poland, Czech), Slovenian, Austrian, Hungary, Belarusian ndi German), 44,45 ndipo, ku United States ndi Argentina, posachedwapa adatanthauzidwa ngati "mtundu wa germline wobwerezabwereza" mwa odwala aku Italy omwe ali ndi BC ndi OC. Mtundu wa 34c.514del udapezeka kale mwa odwala 8 a khansa ya m'mawere ochokera kumpoto kwa Sicily ku Palermo ndi Messina. Chosangalatsa n'chakuti, ngakhale Incorvaia et al. anapeza c.3253dupA m'mabanja ena ku Catania.28 Ma PV a BRCA2 omwe ndi odziwika kwambiri ndi c.428dup, c.5851_5854delAGTT ndi intronic variant c.8487+1G>A, omwe anenedwa mwatsatanetsatane 28 mwa wodwala ku Palermo yemwe ali ndi c.428dup, c.5851_5854delAGTT PV idawonedwa m'mabanja kumpoto chakumadzulo kwa Sicily, makamaka m'madera a Trapani ndi Palermo, pomwe c.5851_5854delAGTT PV idawonedwa m'mabanja kumpoto chakumadzulo kwa Sicily. Mtundu wa 8487+1G>A unali wofala kwambiri mwa anthu ochokera ku Messina, Palermo, ndi Caltanissetta.28 Rebbeck et al. adafotokoza kale za kusintha kwa c.5851_5854delAGTT ku Colombia.37 PV ina ya BRCA2, c.631+1G>A, yapezeka mwa odwala a BC ndi OC ochokera ku Sicily (Agrigento, Siracusa ndi Ragusa).28 Chodziwika bwino n'chakuti, tawona kukhalapo kwa mitundu iwiri ya BRCA2 (BRCA2 c.631G>A ndi c.7008-2A>T) mwa wodwala yemweyo, yemwe timaganiza kuti wapatulidwa mu cis mode, monga momwe zidanenedwera kale.34,46 Kusintha kumeneku kwa BRCA2 kumachitika kawirikawiri m'chigawo cha Italy ndipo kwapezeka kuti kumayambitsa ma codon osakhazikika msanga, zomwe zimakhudza kugawanika kwa messenger RNA ndikupangitsa kuti puloteni ya BRCA2 ilephere.47,48
Tinajambulanso ma PV a BRCA1 ndi BRCA2 m'madera oyerekeza a OCCR ndi BCCR a madera a mapuloteni ndi majini. Madera awa adafotokozedwa ndi Rebbeck et al. ngati madera omwe ali pachiwopsezo chotenga khansa ya ovarian ndi bere, motsatana.49 Komabe, umboni wokhudzana ndi mgwirizano pakati pa malo a mitundu yosiyanasiyana ya majeremusi ndi chiopsezo cha khansa ya m'mawere kapena ovarian ukadali wotsutsana.28,50-52 Mu anthu athu, ma PV a BRCA1 anali ambiri m'dera la BCCR, pomwe ma PV a BRCA2 anali ambiri m'dera la OCCR. Komabe, sitinathe kupeza mgwirizano uliwonse pakati pa madera oyerekeza a OCCR ndi BCCR ndi mawonekedwe a BC. Izi zitha kukhala chifukwa cha kuchuluka kochepa kwa odwala omwe ali ndi kusintha kwa BRCA1/2. Kuchokera pamalingaliro a chigawo cha mapuloteni, ma PV a BRCA1 amagawidwa m'mapuloteni onse, ndipo kusintha kwa BRCA2 kumapezeka makamaka m'dera lobwerezabwereza la BRC.
Pomaliza, tinagwirizanitsa zizindikiro za BC clinicopathological ndi BRCA1/2 PV. Chifukwa cha chiwerengero chochepa cha odwala omwe adaphatikizidwa, tinapeza mgwirizano wofunikira pakati pa Ki-67 ndi mtundu wa chotupa. Ngakhale kuwunika ndi kutanthauzira kwa Ki-67 kukupitilirabe kutsutsana, ndikotsimikiza kuti kuchuluka kwa anthu omwe akuchulukirachulukira kumalumikizidwa ndi chiopsezo chowonjezeka cha kubwereranso kwa matenda ndikuchepa kwa kupulumuka. Mpaka pano, malire osiyanitsa pakati pa Ki-67 "yokwera" ndi "yotsika" ndi 20%. Komabe, malire awa sagwira ntchito kwa odwala athu osinthika a BRCA1/2, omwe ali ndi mtengo wapakati wa Ki-67 wa 25%. Kuchuluka kwa Ki-67 kumeneku kumatha kufotokozedwa ndi kufalikira kwa magulu athu a luminal B ndi TNBC, omwe anali ndi zotupa zochepa za luminal A. Komabe, umboni wina ukuwoneka kuti ukusonyeza kuti kudula kwa Ki-67 kokwera (25-30%) kumatha kugawa bwino odwala malinga ndi zomwe akuyembekezera.53,54 Kuchokera ku zotsatira za kusanthula kwathu, kulumikizana kwakukulu sikudabwitsa. Kumachitika pakati pa Ki-67 yokwera ndi magiredi ndi kukhalapo kwa BRCA1 PV. Ndipotu, zotupa zokhudzana ndi BRCA1 ndizofala kwambiri ku TNBC ndipo zimakhala ndi zizindikiro zowopsa kwambiri.16,17
Pomaliza, kafukufukuyu akupereka lipoti la momwe BRCA1/2 imasinthira m'thupi m'gulu la BC kuchokera kum'mawa kwa Sicily. Ponseponse, zomwe tapeza zikugwirizana ndi umboni womwe ulipo kale, ponse pawiri pankhani ya kuchuluka kwa masinthidwe a thupi komanso zizindikiro za matenda mu BC. Kafukufuku wochulukirapo mwa anthu ambiri odwala BRCA1/2-mutant BC, monga kugwiritsa ntchito multigenome expanded mutational analysis, akuyenera kuwunika kupezeka kwa ma PV omwe ndi osiyana komanso ochepa kuposa BRCA1/2. Izi zithandiza kuzindikira ndi kuyang'anira bwino kuchuluka kwa anthu omwe ali pachiwopsezo chachikulu cha khansa chifukwa cha kusintha kwa majini.
Tatsimikiza kuti odwala adasaina chilolezo chodziwitsa kuti atulutse zitsanzo zawo za chotupa mosadziwika kuti afufuze. Odwala onse adasaina chilolezo chodziwitsa malinga ndi Chikalata cha Helsinki. Malinga ndi mfundo za AOU Policlinico "G.Rodolico - S.Marco", kafukufukuyu sanaloledwe kuunikanso makhalidwe abwino chifukwa kusanthula kwa BRCA1/2 kunachitika malinga ndi machitidwe azachipatala ndipo odwala onse adapereka chilolezo chodziwitsa. Odwala amavomerezanso kugwiritsa ntchito deta yawo pazifukwa zofufuzira.
Tikuthokoza Pulofesa Paolo Vigneri chifukwa cha thandizo lake posamalira odwala khansa ya m'mawere monga momwe Komiti ya Makhalidwe Abwino idapempha.
Federica Martorana akunena za ulemu wochokera kwa Istituto Gentili, Eli Lilly, Novartis, Pfizer. Olemba enawo sananene kuti pali kusagwirizana kulikonse pa ntchito imeneyi.
1. Sung H, Ferlay J, Siegel RL, ndi ena. Ziwerengero za Khansa Padziko Lonse 2020: GLOBOCAN imawerengera kuchuluka ndi kufa kwa khansa 36 m'maiko 185 padziko lonse lapansi. CA Khansa J Clin.2021;71(3):209-249.doi: 10.3322/caac.21660